The molecular characterization of stress responses in prokaryotes and eukaryotes has provided important new insights into the mechanisms of gene regulation. Two well characterized stress responses, the heat shock response and the bacterial 'SOS response' have provided model systems for understanding how multiple genetic elements can be transcriptionally activated in response to environmental signals. The DDR genes of the unicellular eukaryote Saccharomyces cerevisiae are a set of genes which are transcriptionally responsive to DNA damage. Moreover, three of these genes (DDRA2, DDR48 and UBI4) are also induced by heat shock in yeast. The molecular features of transcriptional regulation of the DDR genes will be investigated in order to: i) identify the cis-acting sequences that are necessary and sufficient for DNA damage stress and heat shock induction of transcription; ii) purify the trans-acting factors that are responsible for modulating transcription following stress; and iii) isolate mutations in genes that participate in the stress response pathways and to clone these elements. Promoter fusions will be constructed using upstream sequences of the DDR genes which, based upon deletion analysis are important for stress regulation, and a heterologous CYC-LACZ reporter gene in order to precisely define those DNA sequences that confer both DNA damage and heat shock responsiveness. These promoter fusion constructs will be used to isolate mutations that alter the regulation of the CYC-LACZ reporter gene following DNA damage or heat stress. Gel retardation experiments will be employed to identify proteins which bind to these important regulatory regions. These experiments will provide answers to the following questions: i) Do DNA damage and heat shock stress regulate DDR gene expression through common upstream control elements? ii) Are identical trans-acting factors responsible for both DNA damage and heat shock responses? iii) Are the DDR genes coordinately regulated? and iv) How many genes participate in the stress response regulation to the DDR genes? These studies will provide important insights into the molecular mechanisms of gene regulation in eukaryotes and contribute to our knowledge of the molecular responses of cells to mutagen/carcinogen exposure.